The outbreak of COVID-19 caused a worldwide public health crisis. Large-scale population screening is an effective means to control the spread of COVID-19. Reverse transcription− polymerase chain reaction (RT-qPCR) and serology assays are the most available techniques for SARS-CoV-2 detection; however, they suffer from either less sensitivity and accuracy or low instrument accessibility for screening. To balance the sensitivity, specificity, and test availability, here, we developed enhanced colorimetry, which is termed as a magnetic pull-down-assisted colorimetric method based on the CRISPR/Cas12a system (M-CDC), for SARS-CoV-2 detection. By this method, SARS-CoV-2 RNA from synthetic sequences and cultured viruses can be detected by the naked eye based on gold nanoparticle (AuNP) probes, with a detection limit of 50 RNA copies per reaction. With CRISPR/Cas12a-assisted detection, SARS-CoV-2 can be specifically distinguished from other closely related viruses. M-CDC was further used to analyze 41 clinical samples, whose performance was 95.12%, consistent with that of an approved Clinical RT-qPCR Diagnosis kit. The developed M-CDC method is not dependent on sophisticated instruments, which makes it potentially valuable to be applied for SARS-CoV-2 screening under poor conditions.
Many plasma membrane (PM) functions depend on the cholesterol concentration in the PM in strikingly nonlinear, cooperative ways: fully functional in the presence of physiological cholesterol levels (35~45 mol%), and nonfunctional below 25 mol% cholesterol; namely, still in the presence of high concentrations of cholesterol. This suggests the involvement of cholesterol-based complexes/domains formed cooperatively. In this review, by examining the results obtained by using fluorescent lipid analogs and avoiding the trap of circular logic, often found in the raft literature, we point out the basic similarities of liquid-ordered (Lo)-phase domains in giant unilamellar vesicles, Lo-phase-like domains formed at lower temperatures in giant plasma membrane vesicles, and detergent-resistant membranes: these domains are formed by cooperative interactions of cholesterol, saturated acyl chains, and unsaturated acyl chains. The literature contains good evidence, showing that the domains formed by the same basic cooperative molecular interactions exist and play essential roles in signal transduction in the PM. Therefore, as a working definition, we propose that raft domains in the PM are liquidlike molecular complexes/domains formed by cooperative interactions of cholesterol with saturated acyl chains as well as unsaturated acyl chains, due to saturated acyl chains' weak multiple accommodating interactions with cholesterol and cholesterol's low miscibility with unsaturated acyl chains and TM proteins. Molecules move within raft domains and exchange with those in the bulk PM. We provide a logically established collection of fluorescent lipid probes that preferentially partition into raft and non-raft domains, as defined here, in the PM.
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